title: GROα regulates human embryonic stem cell self-renewal or adoption of a neuronal fate.
abstract: Previously we reported that feeders formed from human placental fibroblasts (hPFs) support derivation and long-term self-renewal of human embryonic stem cells (hESCs) under serum-free conditions. Here, we show, using antibody array and ELISA platforms, that hPFs secrete ∼6-fold higher amounts of the CXC-type chemokine, GROα, than IMR 90, a human lung fibroblast line, which does not support hESC growth. Furthermore, immunocytochemistry and immunoblot approaches revealed that hESCs express CXCR, a GROα receptor. We used this information to develop defined culture medium for feeder-free propagation of hESCs in an undifferentiated state. Cells passaged as small aggregates and maintained in the GROα-containing medium had a normal karyotype, expressed pluripotency markers, and exhibited apical-basal polarity, i.e., had the defining features of pluripotent hESCs. They also differentiated into the three primary (embryonic) germ layers and formed teratomas in immunocompromised mice. hESCs cultured as single cells in the GROα-containing medium also had a normal karyotype, but they downregulated markers of pluripotency, lost apical-basal polarity, and expressed markers that are indicative of the early stages of neuronal differentiation-βIII tubulin, vimentin, radial glial protein, and nestin. These data support our hypothesis that establishing and maintaining cell polarity is essential for the long-term propagation of hESCs in an undifferentiated state and that disruption of cell-cell contacts can trigger adoption of a neuronal fate.
authors.affiliation: SLL Sciences, StemLifeLine, Inc., San Carlos, CA 94070, USA.
authors.forename: Ana
authors.initials: A
authors.lastname: Krtolica
authors.forename: Nick
authors.initials: N
authors.lastname: Larocque
authors.forename: Olga
authors.initials: O
authors.lastname: Genbacev
authors.forename: Dusko
authors.initials: D
authors.lastname: Ilic
authors.forename: Jean-Philippe
authors.initials: JP
authors.lastname: Coppe
authors.forename: Christopher K
authors.initials: CK
authors.lastname: Patil
authors.forename: Tamara
authors.initials: T
authors.lastname: Zdravkovic
authors.forename: Michael
authors.initials: M
authors.lastname: McMaster
authors.forename: Judith
authors.initials: J
authors.lastname: Campisi
authors.forename: Susan J
authors.initials: SJ
authors.lastname: Fisher
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D016764:Cell Polarity; D054360:Chemokine CXCL1 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D003470:Culture Media; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D051379:Mice; D055495:Neurogenesis; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D010920:Placenta / Q000166:cytology / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011247:Pregnancy; D054387:Receptors, CXCR / Q000235:genetics / Q000378:metabolism
chemical_list: D054360:Chemokine CXCL1; D003470:Culture Media; D054387:Receptors, CXCR